The long term goal of the proposed research is to develop an understanding at the cellular and molecular levels, of the events relating to calcium metabolism in the kidney. Of particular interest are those events involved in the regulation of 25- hydroxyvitamin D3 (25-OH-D3) to 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). Specifically the goals are to: (1) Continue studies on the regulation of 25-OH-D3 metabolism in chick kidney cell cultures by examining in detail the extent to which protein kinase C is involved in this process. These studies will contribute not only to our understanding of this intracellular second messenger system in the kidney as it relates to 25-Oh-D3 metabolism, but in other steroidogenic tissues as well. (2) Determine the sequence of the chick and human mRNA for the cytochrome P-450 component of 25-Oh-D3-1-hydroxylase (cytochrome P-450-1 alpha), the enzyme which catalyzes for formation of 1,25(OH)2D3, and use this information to elucidate the position of this protein in the supergene family of cytochrome P-450's. (3) Determine how the mRNA for the cytochrome P-450-1 alpha is regulated by 1,25(OH)2D3 in vivo and by 1,25(OH)2D3 and other known regulators of 25-OH-D3-1-hydroxylase activity in chick kidney cell cultures. (4) Isolate and characterize, chemically and biologically, the newly discovered 45 kDa protein chick kidney mitochondrial protein which is induced by vitamin D deficiency. Determine the relationship of this protein to vitamin D and/or calcium metabolism. these studies will fill the gaps in our knowledge about cytochrome P-450-1 alpha and will open up possibilities leading to increased understanding of diseases related to calcium metabolism, for example, vitamin D dependent rickets, Type I, at the molecular level.